Volume 18 Issue 3, March 2011

Glutamate receptor ion channels use the free energy of ligand binding to trigger ion channel activation and desensitization. In this issue, an analysis of all-atom molecular dynamics simulations dissects the binding process, reveals a substantial gain in free energy produced by domain closure for agonists and reports unique energy landscapes for individual ligands.

Genetic selection uncovers a novel periplasmic chaperone called Spy, whose expression is strongly induced by conditions that cause protein unfolding. Spy can suppress protein aggregation and promote refolding in an ATP-independent manner in vitro. The crystal structure of Spy reveals a thin, cradle-shaped dimer that might shield aggregation-prone regions exposed in unfolded substrates.

Xrn1 is a 5′-3′ exoribonuclease that is implicated in a range of regulated mRNA decay processes in eukaryotes. The structure of Xrn1 is now presented, indicating that it unexpectedly carries repeated structural domains with similarities to the Tudor, Chromo and other domains. These may stabilize the active site and form a platform for protein recruitment.

FliJ interacts with many flagellar proteins involved in protein export, but its function was unclear. Now the crystal structure of FliJ shows it is a homolog of the F1-ATPase γ-subunit. Binding of FliI to FliJ promotes FliI hexameric ring formation, suggesting that flagellar type III protein export systems and F1-ATP synthase use a similar mechanism.

Binding of a neurotransmitter to the ligand-binding domain of glutamate receptors results in conformational changes and opening of the channel. The relationship between ligand binding and activation is not clear, and now the energetics of ligand binding are dissected using computational methods.

The structural and thermodynamic changes caused by substrate binding were followed using a combination of techniques. The approach reveals that allosteric communication between the two active sites of a homodimeric enzyme occurs through two opposing mechanisms. One follows a classical paradigm and the other involves a recently-proposed interplay of folding and binding.

Hsc70 disassembles the coats of clathrin-coated vesicles, remodels a number of other protein complexes, and facilitates protein folding. The dynamics of clathrin uncoating promoted by Hsc70 have now been monitored with single-particle fluorescence imaging. The results suggest that disassembly is driven by trapping of small conformational fluctuations.

The C. elegans microRNA let-7 regulates developmental progression, and human let-7 has been implicated in disease. Examination of endogenous let-7 expression in C. elegans now reveals a complex regulation process whereby pri-let-7 is regulated transcriptionally with oscillations during each larval stage, and co-transcriptionally by LIN-28 which prevents mature microRNA accumulation during early larval stages.

CP29 is one of the minor light-harvesting complexes from plant photosystem II; it absorbs and transfers solar energy for photosynthesis. Now the crystal structure of spinach CP29 is solved, revealing a network of pigments and offering insights into energy transfer pathways.

Ribonucleotide reductase is essential to maintain the cellular pools of dNTPs, and its activity is controlled allosterically by ATP (activator) and dATP (inhibitor). Now crystal and EM structures of human and yeast ribonucleotide reductase 1 in complex with different nucleotides, together with mutagenesis and functional analysis, reveal how dATP binding induces hexamerization and consequence inhibition of the enzyme.

Exportin-5 is involved in microRNA precursor nuclear export. Dicer mRNA is now found to compete with pre-miRNAs for exportin-5 binding, suggesting that cytoplasmic levels of Dicer and its pre-miRNA substrate engage in a feedback loop. Analysis of adenoviral RNA indicates that it can also compete for exportin-5, suggesting that this might be a mechanism by which adenovirus can interfere with host miRNA processing.

The formation of aggregates of polyglutamine (polyQ) sequences is initiated by nucleated growth polymerization. New results show that over a short repeat length range from Q26 to Q23 the size of the nucleus changes from monomeric to dimeric to tetrameric, suggesting aggregation nucleus size has a role in pathogenicity.

Alternative splicing is a regulated process that increases the coding potential of the genome by including or excluding exons. The covalent histone modification H3K9me3, as well as HP1γ,which binds this modification, are now shown to participate in a mechanism that tags a subset of variant exons for inclusion.

Hsp70 chaperones interact with many proteins through a substrate-trapping mechanism that requires ATP. Hsp70s have a lid over the substrate-binding cleft, whose function is controversial. Using cysteine cross-linking and EPR in the Escherichia coli Hsp70 DnaK, it is now shown that helix B of the lid subdomain can adopt three major conformational states and that it does not necessarily close over bound substrates, allowing binding of natively folded states.

RNA Pol III transcribes a number of non-coding RNA types. The crystal structure of hRPC62, part of a ternary subcomplex of RNA Pol III needed for specific initiation, is now described along with its interactions within the subcomplex. hRPC62 shares structural features with the general Pol II transcription factor TFIIEα, indicating functional similarities between components needed for promoter-specific initiation in the two systems.

Riboswitches are RNA elements that provide feedback regulation of metabolic genes upon ligand binding. The glmS riboswitch is also a ribozyme, self-cleaving upon glucosamine-6-phosphate binding. By following intracellular self-cleavage under different growth conditions, it is now found that this riboswitch also responds to inhibitory metabolites, arguing that it integrates multiple signals in response to the metabolic status of the cell.

Knowledge of the molecular mechanisms of activation of Clostridium difficile toxins will significantly enhance the ability to design specific anti-virulence therapies. Using activity-based chemical probes in combination with other techniques, this study reveals mechanistic insights into how inositol hexakisphosphate binding at the active site of the cysteine protease domain shifts the conformational equilibrium towards an active conformer.

Insulin secretion from pancreatic b cells is tightly regulated. Using chromosome conformation capture techniques, it is now found that the insulin promoter locus forms a long-range glucose-regulated interaction with SYT8, linked here to insulin secretion. SYT8 expression is affected concordantly with changes in insulin expression, suggesting a mechanism whereby insulin expression and secretion might be coordinated.

MutS scans DNA for mismatches; once a lesion is recognized, MutS initiates a cascade of events that ultimately results in mismatch excision and repair. Now the behavior of MutS on DNA is studied using smFRET, revealing that mismatch recognition triggers ATP binding that switches MutS from a transient scanning clamp to a very stable sliding clamp.

Crystal structures of the protease domain of human HtrA1, with or without its PDZ domain, reveal that this enzyme's activity is controlled by substrate binding to its active site, via an induced-fit mechanism.

The signal sequence on nascent peptides is recognized by the signal recognition particle (SRP), with subsequent targeting of the SRP-ribosomal nascent complex to the membrane. The structure of a signal sequence bound to the core SRP is presented, revealing structural changes in the SRP upon signal sequence binding.

Rhodopsin is a G protein–coupled receptor (GPCR) involved in biological signaling. How the structure of the ligand is connected with larger-scale functional protein dynamics has remained elusive. Solid-state NMR relaxation shows that localized motions of the ligand retinal lead to collective fluctuations of transmembrane helices in the activation mechanism of the photoreceptor.

The lack of sequence-specificity of DNA replication origins in most eukaryotes poses a challenge for its identification. Through BrdU labeling followed by high-throughput sequencing, supported by ChIP-chip to locate ORC1 and CDC6, the origins of replication in Arabidopsis thaliana are now mapped and their contexts analyzed.